A calcium-dependent galactose-binding lectin from the tunicate Polyandrocarpa misakiensis. Isolation, characterization, and amino acid sequence.
A lectin was isolated from the homogenate of the tunicate Polyandrocarpa misakiensis by heat treatment, ammonium sulfate fractionation, gel filtration, and high-performance ion-exchange chromatography. Analytical gel filtration on Superose 12 and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the lectin is a monomeric protein with a molecular mass of approximately 15 kDa. The lectin bound to an immobilized D-galactose column in the presence of calcium ion with a threshold of 500 microM and eluted completely with 5 mM EDTA. It did not bind to an immobilized D-mannose or N-acetyl-D-galactosamine column. Thus, Polyandrocarpa lectin was found to be a calcium-dependent galactose-binding lectin. The complete amino acid sequence of Polyandrocarpa lectin was determined by automated or manual Edman sequencing of the peptides derived by digestion with trypsin, endoproteinase Asp-N, Staphylococcus aureus V8 protease, and pepsin. It is composed of 125 residues, contains no carbohydrate group, and has a calculated molecular mass of 14,034 Da. The lectin contains four half-cystines, and Cys-21 and Cys-119 and also Cys-96 and Cys-111 form intrachain disulfide bridges, respectively. The amino acid sequence of Polyandrocarpa lectin shows about 20-30% homology with those of fly, barnacle, sea urchin, and several vertebrate lectins that belong to C-type lectin (Drickamer, K. (1988) J. Biol. Chem. 263, 9557-9560). Although the physiological role of Polyandrocarpa lectin is not clear, preliminary experiments suggest that the lectin may be related to defense mechanisms because it has a strong antibacterial activity.
Suzuki
T
T
Department of Biology, Faculty of Science, Kochi University, Japan.
Takagi
T
T
Furukohri
T
T
Kawamura
K
K
Nakauchi
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002845:Chromatography; D005690:Galactose / Q000378:metabolism*; D037102:Lectins / Q000145:classification / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D014561:Urochordata / Q000032:analysis*
D037102:Lectins; D002118:Calcium; D005690:Galactose